Formulation containing anti-inflammatory androstane derivative

ABSTRACT

There is provided a pharmaceutical formulation comprising an aqueous suspension of particulate compound of formula (I)  
                 
 
     or a solvate thereof.

[0001] This application is a Continuation-in-part of a U.S. Pat. No. 35USC 371 patent application, Ser. No. 09/958050 filed on Oct. 2, 2001,for which an International Patent Application No. PCT.GB01.03495 filedAug. 3, 2001, which claims priority to United Kingdom Patent ApplicationNo. GB 0019172.6 filed Aug. 5, 2000.

[0002] The present invention relates to pharmaceutical formulationscontaining an anti-inflammatory and anti-allergic compound of theandrostane series and to processes for their preparation. The presentinvention also relates to therapeutic uses thereof, particularly for thetreatment of inflammatory and allergic conditions.

[0003] Glucocorticoids which have anti-inflammatory properties are knownand are widely used for the treatment of inflammatory disorders ordiseases such as asthma and rhinitis. For example, U.S. Pat. No.4,335,121 discloses 6α,9α-Difluoro-17α(1-oxopropoxy)-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (known by the generic name of fluticasonepropionate) and derivatives thereof. The use of glucocorticoidsgenerally, and especially in children, has been limited in some quartersby concerns over potential side effects. The side effects that arefeared with glucocorticoids include suppression of theHypothalamic-Pituitary-Adrenal (HPA) axis, effects on bone growth inchildren and on bone density in the elderly, ocular complications(cataract formation and glaucoma) and skin atrophy. Certainglucocorticoid compounds also have complex paths of metabolism whereinthe production of active metabolites may make the pharmacodynamics andpharmacokinetics of such compounds difficult to understand. Whilst themodern glucocorticoids are very much safer than those originallyintroduced, it remains an object of research to produce new moleculesand formulations of old and new molecules which have excellentanti-inflammatory properties, with predictable pharmacokinetic andpharmacodynamic properties, with an attractive side effect profile, andwith a convenient treatment regime.

[0004] We have now identified a novel glucocorticoid compound andformulation thereof which substantially meets these objectives, inparticular one suitable for intranasal administration.

[0005] Many millions of individuals suffer from seasonal and perennialallergic rhinitis worldwide. Symptoms of seasonal and perennial allergicrhinitis include nasal itch, congestion, runny nose, sneezing and wateryeyes. Seasonal allergic rhinitis is commonly known as ‘hay fever’. It iscaused by allergens which are present in the air at specific times ofthe year, for example tree pollen during Spring and Summer. Perennialallergic rhinitis is caused by allergens which are present in theenvironment during the entire year, for example dust mites, mold, mildewand pet dander.

[0006] To formulate an effective pharmaceutical nasal composition, themedicament must be delivered readily to all portions of the nasalcavities (the target tissues) where it performs its pharmacologicalfunction. Additionally, the medicament should remain in contact with thetarget tissues for relatively long periods of time. The longer themedicament remains in contact with the target tissues, the medicamentmust be capable of resisting those forces in the nasal passages thatfunction to remove particles from the nose. Such forces, referred to as‘mucociliary clearance’, are recognised as being extremely effective inremoving particles from the nose in a rapid manner, for example, within10-30 minutes from the time the particles enter the nose.

[0007] Other desired characteristics of a nasal composition are that itmust not contain ingredients which cause the user discomfort, that ithas satisfactory stability and shelf-life properties, and that it doesnot include constituents that are considered to be detrimental to theenvironment, for example ozone depletors. In the case of administrationof glucocorticoids, the potential for any undesirable side effectsshould preferably be minimised.

[0008] Thus, according to one aspect of the invention, there is provideda pharmaceutical formulation comprising an aqueous suspension ofparticulate compound of formula (I)

[0009] or a solvate thereof.

[0010] Preferably, the formulation will contain one or more suspendingagents.

[0011] Preferably, the formulation will contain one or morepreservatives.

[0012] Preferably, the formulation will contain one or more wettingagents.

[0013] Preferably, the formulation will contain one or more isotonicityadjusting agents.

[0014] According to one particular aspect of the present invention weprovide a pharmaceutical formulation which comprises:

[0015] (i) an aqueous suspension of particulate compound of formula (1)or a solvate thereof;

[0016] (ii) one or more suspending agents;

[0017] (iii) one or more preservatives;

[0018] (iv) one or more wetting agents; and

[0019] (v) one or more isotonicity adjusting agents.

[0020] The formulations of the present invention may be stabilised byappropriate selection of pH using hydrochloric acid. Typically, the pHwill be adjusted to between 4.5 and 7.5, preferably between 5.0 and 7.0,especially around 6.5.

[0021] Examples of pharmaceutically acceptable materials which can beused to adjust the pH of the formulation include hydrochloric acid andsodium hydroxide. Preferably, the pH of the formulation will be adjustedusing hydrochloric acid.

[0022] The aqueous component is preferably a high grade quality ofwater, most preferably purified water.

[0023] The active compound of formula (I) or solvate thereof willsuitably have a mass mean diameter (MMD) of less than 20 μm, preferablybetween 0.5-10 μm, especially around 3-5 μm, eg. 2 μm. Particle sizereduction, if necessary, may be achieved eg. by micronisation.Preferably, the particles will be crystalline, prepared for example by aprocess which comprises mixing in a continuous flow cell in the presenceof ultrasonic radiation a flowing solution of compound of formula (I) orsolvate thereof as medicament in a liquid solvent with a flowing liquidantisolvent for said medicament (as described in International PatentApplication PCT/GB99/04368).

[0024] A pharmaceutically effective amount of particulate compound offormula (I) or solvate thereof is present within the formulation, in anamount which is preferably between 0.005% and 1% (w/w), preferablybetween 0.01% and 0.5% (w/w), especially 0.05-0.1% (w/w) based on thetotal weight of the formulation. Typically, 50 μl of suspension willcontain 50 μg of compound of formula (I) or solvate thereof.

[0025] Examples of suspending agents include carboxymethylcellulose,veegum, tragacanth, bentonite, methylcellulose and polyethylene glycols.Preferably, the suspending agent will be microcrystalline cellulose andcarboxy methylcellulose sodium, most preferably used as the brandedproduct Avicel RC591 (which typically contains 87-91% microcrystallinecellulose and 9-13% carboxy methylcellulose sodium). Particulatemicrocrystalline cellulose will preferably have a particle size in therange 1 to 100 μm. We believe that Avicel RC591 acts as a suspendingagent by imparting thixotropic properties to the formulation, whereinthe formulation may become a stable suspension upon being stirred,shaken or otherwise disturbed.

[0026] Preferably, the thixotropic nature of the suspending agent willensure that the formulation assumes a gel like appearance at rest,wherein the particulate medicament is dispersed and suspendedsubstantially uniformly, characterised by a high viscosity value. Oncethe composition is subjected to shear forces, such as those caused byagitation prior to spraying, the viscosity of the formulation willpreferably decrease to such a level to enable it to flow readily throughthe spray device and exit as a spray of fine particles in a mist. Theseparticles will then be capable of infiltrating the mucosal surfaces ofthe anterior regions of the nose (frontal nasal cavities), the frontalsinus, the maxillary sinuses and the turbinates which overlie theconchas of the nasal cavities. Once deposited, the viscosity of theformulation will preferably increase to a sufficient level to assume itsgel-like form and resist being cleared from the nasal passages by theinherent mucocillary forces that are present in the nasal cavities.

[0027] When the formulation of the present invention comprises asuspending agent, it will be desirably added in a suitable amount toachieve this function, preferably the suspending agent will be presentwithin the formulation in an amount of between 0.1 and 5% (w/w),especially 1.5% (w/w), based on the total weight of the formulation.

[0028] For stability purposes, the formulation of the present inventionshould be protected from microbial contamination and growth. Examples ofpharmaceutically acceptable anti-microbial agents that can be used inthe formulation include quaternary ammonium compounds (eg. benzalkoniumchloride, benzethonium chloride, cetrimide and cetylpyridiniumchloride), mercurial agents (eg. phenylmercuric nitrate, phenylmercuricacetate and thimerosal), alcoholic agents (eg. chlorobutanol,phenylethyl alcohol and benzyl alcohol), antibacterial esters (eg.esters of para-hydroxybenzoic acid), chelating agents such as disodiumedetate (EDTA) and other anti-microbial agents such as chlorhexidine,chlorocresol, sorbic acid and its salts and polymyxin.

[0029] Preferably, the preservative will comprise disodium edetate,which will preferably be present within the formulation in an amount ofbetween 0.001 and 1% (w/w), especially around 0.015% (w/w), based on thetotal weight of the formulation.

[0030] Preferably the preservative will comprise potassium sorbate whichwill preferably be present within the formultaion in an amount ofbetween 0.01 and 1% (w/w), especially around 0.015% (w/w) based on thetotal weight of the formulation.

[0031] Preferably, the preservative will comprise benzalkonium chloride(BKC), which will preferably be present within the formulation in anamount of between 0.001 and 1% (w/w), especially around 0.015% (w/w),based on the total weight of the formulation.

[0032] More preferably, the preservative comprises disodium edetate andbenzalkonium chloride or disodium edetate and potassium sorbate,particularly disodium edetate and benzalkonium chloride.

[0033] Formulations, eg nasal formulations which contain a suspendedmedicament (such as a compound of formula (I) or a solvate thereof) willpreferably contain a pharmaceutically acceptable wetting agent whichfunctions to wet the particles of medicament to facilitate dispersionthereof in the aqueous phase of the composition. Preferably, the amountof wetting agent used will not cause foaming of the dispersion duringmixing.

[0034] It will be appreciated that any agent which is effective inwetting the particles and which is pharmaceutically acceptable can beused. Examples of wetting agents that can be used are fatty alcohols,esters and ethers. Preferably, the wetting agent will be a hydrophilic,non-ionic surfactant, most preferably polyoxyethylene (20) sorbitanmonooleate (supplied as the branded product Polysorbate 80).

[0035] Wherein the formulation of the present invention comprises awetting agent, it will be desirably added in a sufficient quantity toachieve this function, preferably the wetting agent will be presentwithin the formulation in an amount of between 0.001 and 0.05% (w/w),especially 0.025% (w/w), based on the total weight of the formulation.

[0036] The presence of an isotonicity adjusting agent is to achieveisotonicity with body fluids eg fluids of the nasal cavity, resulting inreduced levels of irritancy associated with many nasal formulations.Examples of suitable isotonicity adjusting agents are sodium chloride,dextrose and calcium chloride. Preferably, the isotonicity adjustingagent will be dextrose, most preferably used as dextrose anhydrous.

[0037] When the formulation of the present invention comprises anisotonicity adjusting agent it will be desirably added in a sufficientquantity to achieve this function, preferably the isotonicity adjustingagent will be present within the formulation in an amount of between 0.1and 10% (w/w), especially 5.0% w/w, based on the total weight of theformulation.

[0038] The compound of formula (I) or a solvate thereof and formulationsthereof have potentially beneficial anti-inflammatory or anti-allergiceffects, particularly upon topical administration to the nose,demonstrated by, for example, its ability to bind to the glucocorticoidreceptor and to illicit a response via that receptor, with long actingeffect. Hence, formulations according to the invention are useful in thetreatment of inflammatory and/or allergic disorders of the nose,especially in once-per-day therapy.

[0039] Formulations according to the invention may be prepared bycombining the ingredients in water. If necessary the pH may be adjustedas a final step. Formulations so prepared may then be filled into thereceptacle.

[0040] Aqueous formulations of the invention may also be employed forrectal, aural, otic, oral, topical or parenteral administration oradministration by inhalation for the treatment of other localinflammatory conditions (eg dermatitis, asthma, chronic obstructivepulmonary disease (COPD) and the like). For example formulations of theinvention may be administered to the lung by nebulisation. Suchformulations may employ excipients (eg preservatives, buffers and thelike) appropriate for the route of administration.

[0041] The particularly desirable biological properties of the compoundof formula (I) are now explained below:

[0042] Compound or formula (I) undergoes highly efficient hepaticmetabolism to yield the 17-β carboxylic acid (X) as the sole majormetabolite in rat and human in vitro systems. This metabolite has beensynthesised and demonstrated to be >1000 fold less active than theparent compound in in vitro functional glucocorticoid assays.

[0043] This efficient hepatic metabolism is reflected by in vivo data inthe rat, which have demonstrated plasma clearance at a rate approachinghepatic blood flow and an oral bioavailability of <1%, consistent withextensive first-pass metabolism.

[0044] In vitro metabolism studies in human hepatocytes havedemonstrated that compound (I) is metabolised in an identical manner tofluticasone propionate but that conversion of (I) to the inactive acidmetabolite occurs approximately 5-fold more rapidly than withfluticasone propionate. This very efficient hepatic inactivation wouldbe expected to minimise systemic exposure in man leading to an improvedsafety profile.

[0045] Inhaled glucocorticoids are also absorbed through the lung andthis route of absorption makes a significant contribution to systemicexposure. Reduced lung absorption could therefore provide an improvedsafety profile. Studies with compound of formula (I) have shownsignificantly lower exposure to compound of formula (I) than withfluticasone propionate after dry powder delivery to the lungs ofanaesthetised pigs.

[0046] Examples of disease states in which the compound of formula (I)has utility include inflammatory and/or allergic conditions of the nasalpassages such as rhinitis eg seasonal and perennial rhinitis as well asother local inflammatory conditions such as asthma, COPD and dermatitis.

[0047] It will be appreciated by those skilled in the art that referenceherein to treatment extends to prophylaxis as well as the treatment ofestablished conditions.

[0048] Preferable means for applying the formulation of the presentinvention to the nasal passages is by use of a pre-compression pump.Most preferably, the pre-compression pump will be a VP7 modelmanufactured by Valois SA. Such a pump is beneficial as it will ensurethat the formulation is not released until a sufficient force has beenapplied, otherwise smaller doses may be applied. Another advantage ofthe pre-compression pump is that atomisation of the spray is ensured asit will not release the formulation until the threshold pressure foreffectively atomising the spray has been achieved. Typically, the VP7model may be used with a botte capable of holding 10-50 ml of aformulation. Each spray will typically deliver 50-100 μl of such aformulation, therefore, the VP7 model is capable of providing at least100 metered doses.

[0049] A suitable dosing regime for the formulation of the presentinvention when administered to the nose would be for the patient toinhale deeply subsequent to the nasal cavity being cleared. Duringinhalation the formulation would be applied to one nostril while theother is manually compressed. This procedure would then be repeated forthe other nostril.

[0050] Typically, one or two inhalations per nostril would beadministered by the above procedure up to three times each day,preferably once or twice daily, especially once daily.

[0051] It will be appreciated that the above dosing regime should beadjusted according to the patient's age, body weight and/or symptomseverity.

[0052] As mentioned above, formulations comprising a compound of formula(I) or solvate thereof are useful in human or veterinary medicine, inparticular as an anti-inflammatory and anti-allergic agent.

[0053] There is thus provided as a further aspect of the invention aformulation comprising the compound of formula (I) or solvate thereoffor use in human or veterinary medicine, particularly in the treatmentof patients with inflammatory and/or allergic conditions.

[0054] According to another aspect of the invention, there is providedthe use of a formulation comprising the compound of formula (I) orsolvate thereof for the manufacture of a medicament for the treatment ofpatients with inflammatory and/or allergic conditions.

[0055] In a further or alternative aspect, there is provided a methodfor the treatment of a human or animal subject with an inflammatoryand/or allergic condition, which method comprises administering to saidhuman or animal subject an effective amount of a formulation comprisingthe compound of formula (I) or solvate thereof.

[0056] The compound of formula (I) is long-acting, therefore preferablythe compound will be delivered once-per-day and the dose will beselected so that the compound has a therapeutic effect in the treatmentof respiratory disorders (eg rhinitis) over 24 hours or more.

[0057] The pharmaceutical compositions according to the invention mayalso be used in combination with another therapeutically active agent,for example, an anti-histamine or an anti-allergic. The invention thusprovides, in a further aspect, a combination comprising the compound offormiula (I) or solvate thereof together with another therapeuticallyactive agent, for example, an anti-histamine or an anti-allergic.

[0058] Examples of anti-histamines include methapyrilene or loratadine.

[0059] Other suitable combinations include, for example, otheranti-inflammatory agents eg NSAIDs (eg sodium cromoglycate, nedocromilsodium, PDE4 inhibitors, leukotriene antagonists, iNOS inhibitors,tryptase and elastase inhibitors, beta-2 integrin antagonists andadenosine 2a agonists)) or antiinfective agents (eg antibiotics,antivirals).

[0060] Also of particular interest is use of the compound of formula (I)or a solvate thereof in combination with a phosphodiesterase 4 (PDE4)inhibitor eg cilomilast or a salt thereof.

[0061] Further, there is provided a process for the preparation of suchpharmaceutical compositions which comprises mixing the ingredients.

[0062] The individual compounds of such combinations may be administeredeither sequentially in separate pharmaceutical compositions as well assimultaneously in combined pharmaceutical formulations. Preferablyadditional therapeutically active ingredients are suspended in theformulation together with the compound of formula (I) Appropriate dosesof known therapeutic agents will be readily appreciated by those skilledin the art.

[0063] A process for preparing a compound of formula (I) comprisesalkylation of a thioacid of formula (II)

[0064] or a salt thereof.

[0065] In this process the compound of formula (II) may be reacted witha compound of formula FCH₂L wherein L represents a leaving group (eg ahalogen atom, a mesyl or tosyl group or the like) for example, anappropriate fluoromethyl halide under standard conditions. Preferably,the fluoromethyl halide reagent is bromofluoromethane. Preferably thecompound of formula (II) is employed as a salt, particularly the saltwith diisopropylethylamine.

[0066] In a preferred process for preparing the compound of formula (I),the compound of formula (II) or a salt thereof is treated withbromofluoromethane optionally in the presence of a phase transfercatalyst. A preferred solvent is methylacetate, or more preferablyethylacetate, optionally in the presence of water. The presence of waterimproves solubility of both starting material and product and the use ofa phase transfer catalyst results in an increased rate of reaction.Examples of phase transfer catalysts that may be employed include (butare not restricted to) tetrabutylammonium bromide, tetrabutylammoniumchloride, benzyltributylammonium bromide, benzyltributylammoniumchloride, benzyltriethylammonium bromide, methyltributylammoniumchloride and methyltrioctylammonium chloride. THF has also successfullybeen employed as solvent for the reaction wherein the presence of aphase transfer catalyst again provides a significantly faster reactionrate. Preferably the product present in an organic phase is washedfirstly with aqueous acid eg dilute HCl in order to remove aminecompounds such as triethylamine and diisopropylethylamine and then withaqueous base eg sodium bicarbonate in order to remove any unreactedprecursor compound of formula (II).

[0067] Compound of formula (I) in unsolvated form may be prepared by aprocess comprising:

[0068] (a) Crystallising the compound of formula (I) in the presence ofa non-solvating solvent such as ethanol, methanol, water, ethyl acetate,toluene, methylisobutylketone or mixtures thereof; or

[0069] (b) Desolvating a compound of formula (I) in solvated form (eg inthe form of a solvate with acetone, isopropanol, methylethylketone, DMFor tetrahydrofuran) eg by heating.

[0070] In step (b) the desolvation will generally be performed at atemperature exceeding 50° C. preferably at a temperature exceeding 100°C. Generally heating will be performed under vacuum.

[0071] Compound of formula (I) in unsolvated form has been found toexist in 3 crystalline polymorphic forms, Forms 1, 2 and 3, althoughForm 3 may be an unstable variant of Form 2. The Forms are characterisedby their X-ray diffraction (XRPD) patterns Broadly speaking the Formsare characterised in their XRPD profiles as follows:

[0072] Form 1: Peak at around 18.9 degrees 2Theta

[0073] Form 2: Peaks at around 18.4 amd 21.5 degrees 2Theta

[0074] Form 3: Peaks at around 18.6 and 19.2 degrees 2Theta.

[0075] Forms 1 appears likely to be the thermodynamically most stableform since Forms 2 and 3 are converted into Form 1 on heating.

[0076] A process for preparing a compound of formula (I) as unsolvatedForm 1 polymorph comprises dissolving compound of formula (I) inmethylisobutylketone, ethyl acetate or methyl acetate and producingcompound of formula (I) as unsolvated Form 1 by addition of anon-solvating anti-solvent such as iso-octane or toluene.

[0077] According to a first preferred embodiment of this process thecompound of formula (I) may be dissolved in ethyl acetate and compoundof formula (I) as unsolvated Form 1 polymorph may be obtained byaddition of toluene as anti-solvent. In order to improve the yield,preferably the ethyl acetate solution is hot and once the toluene hasbeen added the mixture is distilled to reduce the content of ethylacetate.

[0078] According to a second preferred embodiment of this process thecompound of formula (I) may be dissolved in methylisobutylketone andcompound of formula (I) as unsolvated Form 1 polymorph may be obtainedby addition of isooctane as anti-solvent.

[0079] Compound of formula (I) in solvated form may be prepared bycrystallising the compound of formula (I) from a solvating solvent suchas acetone or tetrahydrofuran (THF).

[0080] Preferably in processes for preparing formulations of theinvention, the compound of formula (I) will be employed in unsolvatedform, typically unsolvated Form 1.

[0081] Compounds of formula (II) may be prepared from the corresponding17α-hydroxyl derivative of formula (III):

[0082] using for example, the methodology described by G. H. Phillippset al., (1994) Journal of Medicinal Chemistry, 37, 3717-3729. Forexample the step typically comprises the addition of a reagent suitablefor performing the esterification eg an activated derivative of 2-furoicacid such as an activated ester or preferably a 2-furoyl halide eg2-furoyl chloride (employed in at least 2 times molar quantity relativeto the compound of formula (III)) in the presence of an organic base egtriethylamine. The second mole of 2-furoyl chloride reacts with thethioacid moiety in the compound of formula (III) and needs to be removedeg by reaction with an amine such as diethylamine.

[0083] This method suffers disadvantages, however, in that the resultantcompound of formula (II) is not readily purified of contamination withthe by-product 2-furoyldiethylamide. We have therefore invented severalimproved processes for performing this conversion.

[0084] In a first such improved process we have discovered that by usinga more polar amine such as diethanolamine, a more water solubleby-product is obtained (in this case 2-furoyldiethanolamide) whichpermits compound of formula (II) or a salt thereof to be produced inhigh purity since the by-product can efficiently be removed by waterwashing.

[0085] Thus we provide a process for preparing a compound of formula(II) which comprises:

[0086] (a) reacting a compound of formula (III) with an activatedderivative of 2-furoic acid as in an amount of at least 2 moles of theactivated derivative per mole of compound of formula (III) to yield acompound of formula (IIA)

[0087] ; and

[0088] (b) removal of the sulphur-linked 2-furoyl moiety from compoundof formula (IIA) by reaction of the product of step (a) with an organicprimary or secondary amine base capable of forming a water soluble2-furoyl amide.

[0089] In two particularly convenient embodiments of this process wealso provide methods for the efficient purification of the end productwhich comprise either

[0090] (c1)when the product of step (b) is dissolved in a substantiallywater immiscible organic solvent, purifying the compound of formula (II)by washing out the amide by-product from step (b) with an aqueous wash,or

[0091] (c2) when the product of step (b) is dissolved in a watermiscible solvent, purifying the compound of formula (II) by treating theproduct of step (b) with an aqueous medium so as to precipitate out purecompound of formula (II) or a salt thereof.

[0092] In step (a) preferably the activated derivative of 2-furoic acidmay be an activated ester of 2-furoic acid, but is more preferably a2-furoyl halide, especially 2-furoyl chloride. A suitable solvent forthis reaction is ethylacetate or methylacetate (preferablymethylacetate) (when step (c1) may be followed) or acetone (when step(c2) may be followed). Normally an organic base eg triethylamine will bepresent. In step (b) preferably the organic base is diethanolamine. Thebase may suitably be dissolved in a solvent eg methanol. Generally steps(a) and (b) will be performed at reduced temperature eg between 0 and 5°C. In step (c1) the aqueous wash may be water, however the use of brineresults in higher yields and is therefore preferred. In step (c2) theaqueous medium is for example a dilute aqueous acid such as dilute HCl.

[0093] We also provide an alternative process for preparing a compoundof formula (II) which comprises:

[0094] (a) reacting a compound of formula (III) with an activatedderivative of 2-furoic acid in an amount of at least 2 moles ofactivated derivative per mole of compound of formula (III) to yield acompound of formula (IIA); and

[0095] (b) removal of the sulphur-linked 2-furoyl moiety from compoundof formula (IIA) by reaction of the product of step (a) with a furthermole of compound of formula (III) give two moles of compound of formula(II).

[0096] In step (a) preferably the activated derivative of 2-furoic acidmay be an activated ester of 2-furoic acid, but is more preferably a2-furoyl halide, especially 2-furoyl chloride. A suitable solvent forhis step is acetone. Normally an organic base eg triethylamine will bepresent. In step (b) a suitable solvent is DMF or dimethylacetamide.Normally an organic base eg triethylamine will be present. Generallysteps (a) and (b) will be performed at reduced temperature eg between 0and 5° C. The product may be isolated by treatment with acid and washingwith water.

[0097] This aforementioned process is very efficient in that it does notproduce any furoylamide by-product (thus affording inter aliaenvironmental advantages) since the excess mole of furoyl moiety istaken up by reaction with a further mole of compound of formula (II) toform an additional mole of compound of formula (II).

[0098] Further general conditions for the conversion of compound offormula (III) to compound of formula (II) in the two processes justdescribed will be well known to persons skilled in the art.

[0099] According to a preferred set of conditions, however, we havefound that the compound of formula (II) may advantageously be isolatedin the form of a solid crystalline salt. The preferred salt is a saltformed with a base such as triethylamine, 2,4,6-trimethylpyridine,diisopropylethylamine or N-ethylpiperidine. Such salt forms of compoundof formula (II) are more stable, more readily filtered and dried and canbe isolated in higher purity than the free thioacid. The most preferredsalt is the salt formed with diisopropylethylamine. The triethylaminesalt is also of interest.

[0100] Compounds of formula (III) may be prepared in accordance withprocedures described in GB 2088877B. Compounds of formula (III) may alsobe prepared by a process comprising the following steps:

[0101] Step (a) comprises oxidation of a solution containing thecompound of formula (V). Preferably, step (a) will be performed in thepresence of a solvent comprising methanol, water, tetrahydrofuran,dioxan or diethylene glygol dimethylether. So as to enhance yield andthroughput, preferred solvents are methanol, water or tetrahydrofuran,and more preferably are water or tetrahydrofuran, especially water andtetrahydrofuran as solvent. Dioxan and diethylene glygol dimethyletherare also preferred solvents which may optionally (and preferably) beemployed together with water. Preferably, the solvent will be present inan amount of between 3 and 10 vol relative to the amount of the startingmaterial (1 wt.), more preferably between 4 and 6 vol., especially 5vol. Preferably the oxidising agent is present in an amount of 1-9 molarequivalents relative to the amount of the starting material. Forexample, when a 50% w/w aqueous solution of periodic acid is employed,the oxidising agent may be present in an amount of between 1.1 and 10wt. relative to the amount of the starting material (1 wt.), morepreferably between 1.1 and 3 wt., especially 1.3 wt. Preferably, theoxidation step will comprise the use of a chemical oxidising agent. Morepreferably, the oxidising agent will be periodic acid or iodic acid or asalt thereof. Most preferably, the oxidising agent will be periodic acidor sodium periodate, especially periodic acid. Alternatively (or inaddition), it will also be appreciated that the oxidation step maycomprise any suitable oxidation reaction, eg one which utilises airand/or oxygen. When the oxidation reaction utilises air and/or oxygen,the solvent used in said reaction will preferably be methanol.Preferably, step (a) will involve incubating the reagents at roomtemperature or a little warmer, say around 25° C. eg for 2 hours. Thecompound of formula (IV) may be isolated by recrystallisation from thereaction mixture by addition of an anti-solvent. A suitable anti-solventfor compound of formula (IV) is water. Surprisingly we have discoveredthat it is highly desirable to control the conditions under which thecompound of formula (IV) is precipitated by addition of anti-solvent egwater. When the recrystallisation is performed using chilled water (egwater/ice mixture at a temperature of 0-5° C.) although betteranti-solvent properties may be expected we have found that thecrystalline product produced is very voluminous, resembles a soft geland is very difficult to filter. Without being limited by theory webelieve that this low density product contains a large amount ofsolvated solvent within the crystal lattice. By contrast when conditionsof around 10° C. or higher are used (eg around ambient temperature) agranular product of a sand like consistency which is very easilyfiltered is produced. Under these conditions, crystallisation typicallycommences after around 1 hour and is typically completed within a fewhours (eg 2 hours). Without being limited by theory we believe that thisgranular product contains little or no solvated solvent within thecrystal lattice.

[0102] Step (b) will typically comprise the addition of a reagentsuitable for converting a carboxylic acid to a carbothioic acid eg usinghydrogen sulphide gas together with a suitable coupling agent egcarbonyldiimidazole (CDI) in the presence of a suitable solvent egdimethylformamide.

[0103] The advantages of the formulation of the compound of formula (I)according to the invention may include the fact that the formulationsdemonstrate excellent anti-inflammatory properties, with predictablepharmacokinetic and pharmacodynamic behaviour, with an attractiveside-effect profile, rapid onset of action, long duration of action, andare compatible with a convenient regime of treatment in human patients,in particular being amendable to once-per day dosing. Further advantagesmay include the fact that the formulation has desirable physical andchemical properties which allow for ready manufacture and storage.

[0104] The following non-limiting Examples illustrate the invention:

EXAMPLES

[0105] General

[0106]¹H-nmr spectra were recorded at 400 MHz and the chemical shiftsare expressed in ppm relative to tetramethylsilane. The followingabbreviations are used to describe the multiplicities of the signals: s(singlet), d (doublet), t (triplet), q (quartet), m (multiplet), dd(doublet of doublets), ddd (doublet of doublet of doublets), dt (doubletof triplets) and b (broad). Biotage refers to prepacked silica gelcartridges containing KP-Sil run on flash 12i chromatography module.LCMS was conducted on a Supelcosil LCABZ+PLUS column (3.3 cm×4.6 mm ID)eluting with 0.1% HCO₂H and 0.01 M ammonium acetate in water (solventA), and 0.05% HCO₂H 5% water in acetonitrile (solvent B), using thefollowing elution gradient 0-0.7 min 0%B, 0.7-4.2 min 100%B, 4.2-5.3 min0%B, 5.3-5.5 min 0%B at a flow rate of 3 ml/min. The mass spectra wererecorded on a Fisons VG Platform spectrometer using electrospraypositive and negative mode (ES+ve and ES−ve).

[0107] Intermediates

[0108] Intermediate 1: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicAcid Diisopropylethylamine Salt

[0109] A stirred suspension of 6α, 9α-difluoro-11β,17α-dihydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioic acid(prepared in accordance with the procedure described in GB 2088877B)(49.5 g) in methylacetate (500 ml) is treated with triethylamine (35 ml)maintaining a reaction temperature in the range 0-5° C. 2-Furoylchloride (25 ml) is added and the mixture stirred at 0-5° C. for 1 hour.A solution of diethanolamine (52.8 g) in methanol (50 ml) is added andthe mixture stirred at 0-5° C. for at least 2 hours. Dilute hydrochloricacid (approx 1 M, 550 ml) is added maintaining a reaction temperaturebelow 15° C. and the mixture stirred at 15° C. The organic phase isseparated and the aqueous phase is back extracted with methyl acetate(2×250 ml). All of the organic phases are combined, washed sequentiallywith brine (5×250 ml) and treated with di-isopropylethylamine (30 ml).The reaction mixture is concentrated by distillation at atmosphericpressure to an approximate volume of 250 ml and cooled to 25-30° C.(crystallisation of the desired product normally occurs duringdistillation/subsequent cooling). Tertiary butyl methyl ether (TBME)(500 ml) is added, the slurry further cooled and aged at 0-5° C. for atleast 10 minutes. The product is filtered off, washed with chilled TBME(2×200 ml) and dried under vacuum at approximately 40-50° C. (75.3 g,98.7%). NMR (CDCl₃) δ: 7.54-7.46 (1H, m), 7.20-7.12 (1H, dd), 7.07-6.99(1H, dd), 6.48-6.41 (2H, m), 6.41-6.32 (1H, dd), 5.51-5.28 (1H, dddd²J_(H-F) 50 Hz), 4.45-4.33(1H, bd), 3.92-3.73 (3H, bm), 3.27-3.14 (2H,q), 2.64-2.12 (5H, m), 1.88-1.71 (2H, m), 1.58-1.15 (3H, s), 1.50-1.38(15H, m), 1.32-1.23 (1H, m), 1.23-1.15 (3H s), 1.09-0.99 (3H, d)

[0110] Intermediate 2: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16αmethyl-3-oxo-androsta-1,4-diene-17β-carbothioicAcid S-fluoromethyl Ester Unsolvated Form 1

[0111] A mobile suspension of Intermediate 1 (12.61 g, 19.8 mmol) inethyl acetate (230 ml) and water (50 ml) is treated with a phasetransfer catalyst (benzyltributylammonium chloride, 10 mol %), cooled to3° C. and treated with bromofluoromethane (1.10 ml, 19.5 mmol, 0.98equivalents), washing in with prechilled (0° C.) ethyl acetate (EtOAc)(20 ml). The suspension is stirred overnight, allowing to warm to 17° C.The aqueous layer is separated and the organic phase is sequentiallywashed with 1 M HCl (50 ml), 1%w/v NaHCO₃ solution (3×50 ml) and water(2×50 ml). The ethylacetate solution is distilled at atmosphericpressure until the distillate reaches a temperature of approximately 73°C. at which point toluene (150 ml) is added. Distillation is continuedat atmospheric pressure until all remaining EtOAc has been removed(approximate distillate temperature 103° C.). The resultant suspensionis cooled and aged at <10° C. and filtered off. The bed is washed withtoluene (2×30 ml) and the product oven dried under vacuum at 60° C. toconstant weight to yield the title compound (8.77 g, 82%) LCMS retentiontime 3.66 min, m/z 539 MH⁺, NMR δ (CDCl₃) includes 7.60 (1H, m),7.18-7.11 (2H, m), 6.52 (1H, dd, J 4.2 Hz), 6.46 (1H, s), 6.41 (1H, dd,J 10, 2 Hz), 5.95 and 5.82 (2H dd, J 51, 9 Hz), 5.48 and 5.35 (1H, 2m),4.48 (1H, m), 3.48 (1H, m), 1.55 (3H, s), 1.16 (3H, s), 1.06 (3H, d, J 7Hz).

[0112] Pharmacological Activity

[0113] In Vitro Pharmacological Activity

[0114] Pharmacological activity was assessed in a functional in vitroassay of glucocorticoid agonist activity which is generally predictiveof anti-inflammatory or anti-allergic activity in vivo.

[0115] For the experiments in this section, compound of formula (I) wasused as unsolvated Form 1 (Intermediate 2).

[0116] The functional assay was based on that described by K. P. Ray etal., Biochem J. (1997), 328, 707-715. A549 cells stably transfected witha reporter gene containing the NF-κB responsive elements from the ELAMgene promoter coupled to sPAP (secreted alkaline phosphatase) weretreated with test compounds at appropriate doses for 1 hour at 37° C.The cells were then stimulated with tumour necrosis factor (TNF, 10ng/ml) for 16 hours, at which time the amount of alkaline phosphataseproduced is measured by a standard colourimetric assay. Dose responsecurves were constructed from which EC₅₀ values were estimated.

[0117] In this test the compound of formula (I) showed an EC₅₀ value of<1 nM.

[0118] The glucocorticoid receptor (GR) can function in at least twodistinct mechanisms, by upregulating gene expression through the directbinding of GR to specific sequences in gene promoters, and bydownregulating gene expression that is being driven by othertranscription factors (such as NFκB or AP-1) through their directinteraction with GR.

[0119] In a variant of the above method, to monitor these functions, tworeporter plasmids have been generated and introduced separately intoA549 human lung epithelial cells by transfection. The first cell linecontains the firefly luciferase reporter gene under the control of asynthetic promoter that specifically responds to activation of thetranscription factor NFκB when stimulated with TNFα. The second cellline contains the renilla luciferase reporter gene under the control ofa synthetic promotor that comprises 3 copies of the consensusglucocorticoid response element, and which responds to directstimulation by glucocorticoids. Simultaneous measurement oftransactivation and transrepression was conducted by mixing the two celllines in a 1:1 ratio in 96 well plate (40,000 cells per well) andgrowing overnight at 37° C. Test compounds were dissolved in DMSO, andadded to the cells at a final DMSO concentration of 0.7%. Afterincubation for 1 h 0.5 ng/ml TNFα (R&D Systems) was added and after afurther 15 hours at 37° C., the levels of firefly and renilla luciferasewere measured using the Packard Firelite kit following themanufacturers' directions. Dose response curves were constructed fromwhich EC₅₀ values were determined. Transactivation Transrepression (GR)(NFκB) ED₅₀ (nM) ED₅₀ (nM) Compound of Formula (I) 0.06 0.20 Metabolite(X) >250 >1000 Fluticasone propionate 0.07 0.16

[0120] In Vivo Pharmacological Activity

[0121] Pharmacological activity in vivo was assessed in an ovalbuminsensitised Brown Norway rat eosinophilia model. This model is designedto mimic allergen induced lung eosinophilia, a major component of lunginflammation in asthma.

[0122] For the experiments in this section, compound of formula (I) wasused as unsolvated Form 1.

[0123] Compound of formula (I) produced dose dependant inhibition oflung eosinophilia in this model after dosing as an intra-tracheal (IT)suspension in saline 30 min prior to ovalbumin challenge. Significantinhibition is achieved after a single dose of 30 μg of compound offormula (I) and the response was significantly (p=0.016) greater thanthat seen with an equivalent dose of fluticasone propionate in the samestudy (69% inhibition with compound of formula (I) vs 41% inhibitionwith fluticasone propionate).

[0124] In a rat model of thymus involution 3 daily IT doses of 100 μg ofcompound (I) induced significantly smaller reductions in thymus weight(p=0.004) than an equivalent dose of fluticasone propionate in the samestudy (67% reduction of thymus weight with compound (I) vs 78% reductionwith fluticasone propionate).

[0125] Taken together these results indicate a superior therapeuticindex for compound (I) compared to fluticasone propionate.

[0126] In vitro Metabolism in Rat and Human Hepatocytes

[0127] Incubation of compound (I) with rat or human hepatocytes showsthe compound to be metabolised in an identical manner to fluticasonepropionate with the 17-β carboxylic acid (X) being the only significantmetabolite produced. Investigation of the rate of appearance of thismetabolite on incubation of compound (I) with human hepatocytes (37° C.,10 μM drug concentration, hepatocytes from 3 subjects, 0.2 and 0.7million cells/mL) shows compound (I) to be metabolised ca. 5-fold morerapidly than fluticasone propionate: 17-β acid metabolite productionSubject Cell density (pmol/h) number (million cells/mL) Compound (I)Fluticasone propionate 1 0.2 48.9 18.8 1 0.7 73.3 35.4 2 0.2 118 9.7 20.7 903 23.7 3 0.2 102 6.6 3 0.7 580 23.9

[0128] Median metabolite production 102-118 pmol/h for compound (I) and18.8-23.0 pmol/h for fluticasone propionate.

[0129] Pharmacokinetics After Intravenous (IV) and Oral Dosing in Rats

[0130] Compound (I) was dosed orally (0.1 mg/kg) and IV (0.1 mg/kg) tomale Wistar Han rats and pharmacokinetic parameters determined. Compound(I) showed negligible oral bioavailability (0.9%) and plasma clearanceof 47.3 mL/min/kg, approaching liver blood flow (plasma clearance offluticasone propionate=45.2 mL/min/kg).

[0131] Pharmacokinetics After Intra-Tracheal Dry Powder Dosing in thePig.

[0132] Anaesthetised pigs (2) were dosed intra-tracheally with ahomogenous mixture of compound (I) (1 mg) and fluticasone propionate (1mg) as a dry powder blend in lactose (10% w/w). Serial blood sampleswere taken for up to 8 h following dosing. Plasma levels of compound (I)and fluticasone propionate were determined following extraction andanalysis using LC-MS/MS methodology, the lower limits of quantitation ofthe methods were 10 and 20 pg/mL for compound (I) and fluticasonepropionate respectively. Using these methods compound (I) wasquantifiable up to 2 hours after dosing and fluticasone propionate wasquantifiable up to 8 hours after dosing. Maximum plasma concentrationswere observed for both compounds within 15 min after dosing. Plasmahalf-life data obtained from IV dosing (0.1 mg/kg) was used to calculateAUC (0-inf) values for compound (I). This compensates for the plasmaprofile of Compound (I) only being defined up to 2 hours after an ITdose and removes any bias due to limited data between compound (I) andfluticasone propionate.

[0133] C_(max) and AUC (0-inf) values show markedly reduced systemicexposure to compound (I) compared to fluticasone propionate: Cmax(pg/mL) AUC (0-inf) (hr · pg/mL) Pig 1 Pig 2 Pig 1 Pig 2 Compound ofFormula (I) 117  81 254 221 Fluticasone propionate 277 218 455 495

[0134] The pharmacokinetic parameters for both compound (I) andfluticasone propionate were the same in the anaesthetised pig followingintravenous administration of a mixture of the two compounds at 0.1mg/kg. The clearance of these two glucocorticoids is similar is thisexperimental pig model.

EXAMPLES Example 1 Nasal formulation containing 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicAcid S-fluoromethyl Ester

[0135] A formulation for intranasal delivery was prepared withingredients as follows: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared according to Intermediate 2,micronised to MMD 3 μm) 0.05% w/w Polysorbate 80 0.025% w/w Avicel RC591 1.5% w/w Dextrose  5.0% w/w BKC 0.015% w/w EDTA 0.015% w/w water to100%

[0136] in a total amount suitable for 120 actuations and the formulationwas filled into a bottle (plastic or glass) fitted with a metering valveadapted to dispense 50 or 100 μl per actuation

[0137] The device was fitted into a nasal actuator (Valois, e.g. VP3,VP7 or VP7D)

[0138] The formulation was prepared following the following protocol:

[0139] Part A

[0140] 1. Dissolve dextrose in purified water

[0141] 2. Dissolve EDTA in dextrose solution

[0142] 3. Add Avicel RC591 while stirring

[0143] 4. Allow suspension to hydrate

[0144] Part B (separately)

[0145] 1. Dissolve polysorbate 80 in purified water at 50-60° C.

[0146] 2. Prepare slurry of drug in Polysorbate 80 solution

[0147] Part C

[0148] 1. Combine suspension of A4 with suspension of B2 and stir

[0149] 2. Add solution of BKC in purifed water and stir

[0150] 3. Adjust pH with 1 N HCl

[0151] 4. Add purified water to correct weight

Example 2 Nasal Formulation Containing 6α,9α-Difluoro-17α-[2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicAcid S-fluoromethyl Ester

[0152] A formulation for intranasal delivery was prepared withingredients as follows: 6α,9α-Difluoro-17α-[(2-furanylcarbonyl)oxy]-11β-hydroxy-16α-methyl-3-oxo-androsta-1,4-diene-17β-carbothioicacid S-fluoromethyl ester (prepared according to Intermediate 2,micronised to MMD 3 μm) 0.1% w/w Polysorbate 80 0.025% w/w Avicel RC591 1.5% w/w Dextrose  5.0% w/w BKC 0.015% w/w EDTA 0.015% w/w water to100%

[0153] in a total amount suitable for 120 actuations and the formulationwas filled into a bottle fitted with a metering valve adapted todispense 50 or 100 μl per actuation. The device was fitted into a nasalactuator (Valois).

[0154] Stability studies on Examples 1 and 2 showed them to be stable upto 3 months at 40° C. (measurements were not taken beyond this time).

[0155] Throughout the specification and the claims which follow, unlessthe context requires otherwise, the word ‘comprise’, and variations suchas ‘comprises’ and ‘comprising’, will be understood to imply theinclusion of a stated integer or step or group of integers but not tothe exclusion of any other integer or step or group of integers orsteps.

[0156] The patents and patent applications described in this applicationare herein incorporated by reference.

1 A pharmaceutical formulation which comprises: an aqueous suspension ofparticulate compound of formula (I)

or a solvate thereof:
 2. A pharmaceutical formulation according to claim1 which comprises: one or more suspending agents.
 3. A pharmaceuticalformulation according to claim 2 wherein the suspending agent ismicrocrystalline cellulose and carboxy methylcellulose sodium.
 4. Apharmaceutical formulation according to claim 2 wherein the suspendingagent is present in an amount of between 0.1 and 5% (w/w), based on thetotal weight of the formulation.
 5. A pharmaceutical formulationaccording to claim 1 which comprises: one or more preservatives.
 6. Apharmaceutical formulation according to claim 5 wherein the preservativecomprises benzalkonium chloride.
 7. A pharmaceutical formulationaccording to claim 6 wherein the benzalkonium chloride is present withinthe formulation in an amount of between 0.001 and 1% (w/w), based on thetotal weight of the formulation.
 8. A pharmaceutical formulationaccording to claim 5 wherein the preservative comprises EDTA.
 9. Apharmaceutical formulation according to claim 6 wherein the preservativealso comprises EDTA.
 10. A pharmaceutical formulation according to anyclaim 1 which comprises: one or more wetting agents.
 11. Apharmaceutical formulation according to claim 10 wherein the wettingagent comprises polyoxyethylene (20) sorbitan monooleate.
 11. Apharmaceutical formulation according to claim 11 wherein thepolyoxyethylene (20) sorbitan monooleate is present within theformulation in an amount of between 0.001 and 0.05% (w/w), based on thetotal weight of the formulation.
 12. A pharmaceutical formulationaccording to claim 1 which comprises: one or more isotonicity adjustingagents.
 13. A pharmaceutical formulation according to claim 12 whereinthe isotonicity adjusting agent comprises dextrose.
 14. A pharmaceuticalformulation according to claim 13 wherein dextrose is present within theformulation in an amount of between 0.1 and 10% (w/w), based on thetotal weight of the formulation.
 15. A pharmaceutical formulationaccording to claim 1 characterised in that it is isotonic with fluids ofthe nasal cavity.
 16. A pharmaceutical formulation to claim 1 which isbuffered to a pH of between 5 and
 7. 17. A pharmaceutical formulationaccording to claim 16 which is buffered using hydrochloric acid.
 18. Apharmaceutical formulation according to claim 1 wherein the compound offormula (I) or solvate thereof is present within the formulation in anamount between 0.005% and 1% (w/w), based on the total weight of theformulation.
 19. A pharmaceutical formulation according to claim 1wherein the compound of formula (I) is used as unsolvated polymorphForm
 1. 20. A pharmaceutical formulation according to claim 1 whichcomprises (i) one or more suspending agents; (ii) one or morepreservatives; (iii) one or more wetting agents; and (iv) one or moreisotonicity adjusting agents
 21. A pharmaceutical formulation accordingto claim 20 wherein the suspending agent is microcrystalline celluloseand carboxy methylcellulose sodium, the preservative is EDTA andbenzalkonium chloride, the wetting agent is polyoxyethylene (20)sorbitan monooleate and the isotonicity adjusting agent is dextrose. 22.A container comprising a pharmaceutical formulation according to claim 1suitable for delivering it in the form of a nasal spray.
 23. A method oftreatment of allergic rhinitis which comprises administering to apatient a pharmaceutically acceptable amount of a formulation accordingto claim
 1. 24. The method according to claim 23 wherein theadministration is once-per-day.